Peripheral and BM cytopenia in O5A^Δ/Δ mice treated twice with 5-FU. (A) Graphical illustration of our hypothesis of how cytopenia is driven by declining niche homeostasis during consecutive periods of stress. (B) Experimental design for analysis of mice after 2 5-FU treatments (DAYS 0 AND 8). Control (CTRL): O5A^+/+, n = 7, 5A^fl/fl, n = 11 and O5A^Δ/Δ, n = 11. (C) Survival of WT (open symbols, combined results of 5A^fl/fl and O5A^+/+ mice) and O5A^Δ/Δ mice. (D-F) Graphs showing WBC, RBC, and hemoglobin (HGB) measured in PB, respectively, at DAYS 8 AND 14 from WT mice (open symbol, combined results of 5A^fl/fl and O5A^+/+ mice) and O5A^Δ/Δ mice (closed symbols). (G) Total BM cell number 14 days after the first 5-FU treatment in CTRL (n = 9) and O5A^Δ/Δ (n = 5) mice. Analysis of BM from 4 flushed long bones. (H) Representative contour plots of the BM after 2 consecutive 5-FU treatments at day 14. Graphs showing the percentage of LSKs (left) and LT-HSCs (right) in CTRL and O5A^Δ/Δ mice. (I) Representative contour plots of spleen cells from mice treated twice with 5-FU. (J) Graphs showing the absolute cell number of spleen cells (left) and relative number of LSK cells in spleen (right). *P < .05 (2-sided parametric Student’s t test: [C-F]; Mann-Whitney test: [G-H]). The results represent 2 to 3 independent experiments. Data are represented as dots per mouse or cell and the mean ± SEM. Symbol legend shown in Figure 2B.